Chronic limb unloading causes muscle atrophy and conversion of type I (slow twitch) fibers to type II (fast twitch). Atrophy lowers force generating capacity. Conversion of type I to type II fiber: 1) shortens time to peak tension, 2) lowers capacity for oxidative ATP production, 3) lowers capillary density, and 4) lowers metabolic efficiency of cross-bridge cycling. Thus, type II fibers require a greater fraction of their ATP production to come from carbohydrates than fats. Due to the more limited supply of carbohydrates versus fats, there is a metabolic basis for a loss of muscle work capacity accompanying type I to type II fiber conversion. Unfortunately, few data exist regarding how substrate utilization is altered by muscle atrophy, deconditioning, or fiber type conversion. We have used the isolated, perfused, rat hindquarters for these experiments. We have infused [3-13C]lactate or [1-13C]glucose with [U-13C]acetate in Krebs-Henselheit buffer alone and with suspended bovine erythrocytes while electrically stimulating one set of leg muscles via the sciatic nerve. So far, we have demonstrated that the cell can discriminate between pyruvate derived from exogenous [3-13C]lactate and pyruvate derived from exogenous [1-13C]glucose; that acetate incorporation into the citric acid cycle occurs in hypoxic as well as normoxic tissue; that non-oxidative entry of carbon units derived from exogenous [3-13C]lactate and [1-13C]glucose are required from normal citric acid cycle function although their relative contributions are different; that differences in the metabolic capacity between type I (soleus) and type II (gastrocnemius) muscle are reflected by different ratios of oxidative versus non-oxidative entry of label from exogenous [3-13C]lactate and [1-13C]glucose; and that many of these differences reflect acute changes in TCA cycle pool sizes, redox states, and overall muscle oxygen consumptions. This suggests the biochemical basis for a beneficial effect of exercise training as a commonly used countermeasure to the deleterious effects of disuse atrophy associated with microgravity.